Plants and seeds of canola variety scv528587

ABSTRACT

In an embodiment, the invention relates to the seeds, plants, and plant parts of canola variety SCV528587 and to methods for producing a canola plant produced by crossing canola variety SCV528587 with itself or with another canola variety. The invention also relates to methods for producing a canola plant containing in its genetic material one or more transgenes and to the transgenic canola plants and plant parts produced by those methods. This invention also relates to canola varieties or breeding lines and plant parts derived from canola variety SCV528587, to methods for producing other canola varieties, lines or plant parts derived from canola variety SCV528587 and to the canola plants, varieties, and their parts derived from use of those methods. The invention further relates to hybrid canola seeds, plants and plant parts produced by crossing the variety SCV528587 with another canola variety.

BACKGROUND OF THE INVENTION Field of the Invention

The present invention relates to a new and distinctive canola variety,designated SCV528587. All publications cited in this application areherein incorporated by reference.

Description of Related Art

Canola, Brassica napus oleifera annua, is an important and valuablefield crop. Thus, a goal of canola plant breeders is to develop stable,high-yielding canola varieties that are agronomically sound. The reasonsfor this goal are generally to maximize the amount of grain produced onthe land used and to supply food for both animals and humans. The highquality vegetable oil extracted from canola grain is a primary reasonfor canola's commercial value. Thus, in addition to high grain yields,increasing the oil content level in the grain can maximize crop valueper acre. To accomplish these goals, the canola breeder must select anddevelop canola plants that have the traits that result in superiorvarieties.

SUMMARY OF THE INVENTION

Reference now will be made in detail to the embodiments of theinvention, one or more examples of which are set forth below. Eachexample is provided by way of explanation of the invention, not alimitation of the invention. In fact, it will be apparent to thoseskilled in the art that various modifications and variations can be madein the present invention without departing from the scope or spirit ofthe invention. For instance, features illustrated or described as partof one embodiment can be used on another embodiment to yield a stillfurther embodiment. Thus, it is intended that the present inventioncovers such modifications and variations as come within the scope of theappended claims and their equivalents. Other objects, features andaspects of the present invention are disclosed in or are obvious fromthe following detailed description. It is to be understood by one ofordinary skill in the art that the present discussion is a descriptionof exemplary embodiments only and is not intended as limiting thebroader aspects of the present invention.

According to the invention, there is provided a new canola varietydesignated SCV528587. This invention thus relates to the seeds, plants,and/or plant parts of canola variety SCV528587 and to methods forproducing a canola plant produced by crossing canola variety SCV528587with itself or another canola genotype, and the creation of variants bymutagenesis or transformation of canola variety SCV528587. Thus, anymethods using the canola variety SCV528587 are part of this invention,for example, selfing, backcrosses, hybrid production, crosses topopulations, and the like. All plants produced using canola varietySCV528587 as a parent are within the scope of this invention, forexample, canola variety SCV528587 could be used in crosses with other,different, canola plants to produce first generation (F₁) canola hybridseeds and plants with superior characteristics.

In another aspect, the present invention provides for single or multiplegene converted plants of SCV528587. The transferred gene(s) may be adominant or recessive allele. The transferred gene(s) may confer suchtraits as herbicide resistance, insect resistance, resistance forbacterial, fungal, or viral disease, male fertility, male sterility,enhanced nutritional quality, modified fatty acid metabolism, modifiedcarbohydrate metabolism, modified seed yield, modified oil percent,modified protein percent, modified lodging resistance, modifiedglucosinolate content, modified chlorophyll content and industrialusage. The gene may be a naturally occurring gene or a transgeneintroduced through genetic engineering techniques.

In another aspect, the present invention provides regenerable cells foruse in tissue culture of, for example, protoplasts, cells, and plantparts derived from a plant of canola SCV528587. The tissue culture willpreferably be capable of regenerating plants having all of themorphological and physiological characteristics of a plant of canolaSCV528587 and of regenerating plants having substantially the samegenotype as a plant of canola SCV528587. Preferably, the regenerablecells in such tissue cultures will be embryos, spheroplasts,protoplasts, meristematic cells, callus, pollen, leaves, anthers,pistils, cotyledons, roots, root tips, flowers, seeds, pods or stems.Still further, the present invention provides canola plants regeneratedfrom the tissue cultures of the invention.

Yet another aspect of the current invention is a canola plant furthercomprising a single locus conversion. In one embodiment, the canolaplant is defined as further comprising the single locus conversion andotherwise capable of expressing all of the morphological andphysiological characteristics of the canola variety SCV528587. Inparticular embodiments of the invention, the single locus conversion maycomprise a transgenic gene which has been introduced by genetictransformation into the canola variety SCV528587 or a progenitorthereof. In still other embodiments of the invention, the single locusconversion may comprise a dominant or recessive allele. The locusconversion may confer potentially any trait upon the single locusconverted plant, including herbicide resistance, insect resistance,resistance to bacterial, fungal, or viral disease, male fertility orsterility, and improved nutritional quality. In certain embodiments, apotential locus conversion that confers herbicide resistance may conferresistance to herbicides such as, for example, imidazolinone herbicides,sulfonylurea herbicides, triazine herbicides, phenoxy herbicides,cyclohexanedione herbicides, benzonitrile herbicides,4-hydroxyphenylpyruvate dioxygenase-inhibiting herbicides,protoporphyrinogen oxidase-inhibiting herbicides, acetolactatesynthase-inhibiting herbicides, 1-aminocyclopropane-1-carboxylic acidsynthase-inhibiting herbicides, bromoxynil, nicosulfuron,2,4-dichlorophenoxyacetic acid (2,4-D), dicamba, quizalofop-p-ethyl,glyphosate, or glufosinate.

In further embodiments, a single locus conversion comprises a geneticmodification to the genome of canola variety SCV528587. A geneticmodification may comprise, for example, an insertion, deletion, orsubstitution of a nucleotide sequence. In certain embodiments, a singlelocus may comprise one or more genes or intergenic regions integratedinto or mutated at a single locus or may comprise one or more nucleicacid molecules integrated at the single locus. In particularembodiments, a single locus conversion may be generated by genomeediting such as through use of engineered nucleases, as is known in theart. Examples of engineered nucleases include, but are not limited to,Cas endonucleases, zinc finger nucleases (ZFNs), transcriptionactivator-like effector nucleases (TALENs), and engineeredmeganucleases, also known as homing endonucleases. Naturally occurringnucleases can also find use for genome editing. In specific embodiments,endonucleases, both naturally occurring and engineered, may utilize anypolypeptide-, DNA-, or RNA-guided genome editing systems known to theskilled artisan.

In another aspect, the present invention provides a method of plantbreeding wherein the method comprises the steps of a) crossing a plantof canola variety SCV528587 with a second canola plant that comprises atrait to produce F₁ progeny plants; b) selecting at least a firstprogeny plant from step (a) that comprises the trait to produce aselected progeny plant; c) crossing the selected progeny plant from step(b) with a plant of canola variety SCV528587 to produce at least a firstprogeny plant that comprises the trait; and d) repeating steps (b) and(c) with the first progeny plant produced from step (c) used in place ofthe first progeny plant of step (b) during said repeating, wherein steps(b) and (c) are repeated until at least a backcross progeny plant isproduced comprising the trait. The invention still further provides acanola plant produced by this and the foregoing methods.

Definitions

In the description and tables which follow, a number of terms are used.In order to provide a clear and consistent understanding of thespecification and claims, including the scope to be given such terms,the following definitions are provided:

Allele: Any of one or more alternative forms of a genetic locus. In adiploid cell or organism, the two alleles of a given locus occupycorresponding loci on a pair of homologous chromosomes.

Alter: The utilization of up-regulation, down-regulation, or genesilencing.

Anther arrangement: The orientation of the anthers in fully openedflowers can also be useful as an identifying trait. This can range fromintrose (facing inward toward pistil), erect (neither inward notoutward), or extrose (facing outward away from pistil).

Anther dotting: The presence/absence of anther dotting (colored spots onthe tips of anthers) and if present, the percentage of anther dotting onthe tips of anthers in newly opened flowers is also a distinguishingtrait for varieties.

Anther fertility: This is a measure of the amount of pollen produced onthe anthers of a flower. It can range from sterile (such as in femaleparents used for hybrid seed production) to fertile (all anthersshedding).

Backcrossing: A process in which a breeder repeatedly crosses hybridprogeny back to one of the parents, for example, a first generationhybrid F₁ with one of the parental genotypes of the F₁ hybrid.Backcrossing can be used to introduce one or more single locusconversions from one genetic background into another.

Blackleg (Leptosphaeria maculans): Virulent or severe blackleg ofcanola/rapeseed is a fungal canker or dry rot disease of the activelygrowing crop that causes stem girdling and lodging. In heavily infestedcrops, up to 100 percent of the stems may be infected, resulting inmajor yield loss. For purposes of this application, resistance toblackleg is measured using ratings of “R” (resistant), “MR” (mediumresistant), “MS” (moderately susceptible) or “S” (susceptible).

Cell: Cell as used herein includes a plant cell, whether isolated, intissue culture or incorporated in a plant or plant part.

Cotyledon width: The cotyledons are leaf structures that form in thedeveloping seeds of canola which make up the majority of the mature seedof these species. When the seed germinates, the cotyledons are pushedout of the soil by the growing hypocotyls (segment of the seedling stembelow the cotyledons and above the root) and they unfold as the firstphotosynthetic leafs of the plant. The width of the cotyledons varies byvariety and can be classified as narrow, medium, or wide.

Crossing: The mating of two parent plants.

Cross-pollination: Fertilization by the union of two gametes fromdifferent plants.

Elite canola line or variety: A canola line or variety, per se, whichhas been sold commercially.

Elite canola parent line or variety: A canola line or variety which is aparent of a canola hybrid which has been commercially sold.

Emasculate: The removal of plant male sex organs or the inactivation ofthe organs with a cytoplasmic or nuclear genetic factor or a chemicalagent conferring male sterility.

Embryo: The embryo is the small plant contained within a mature seed.

Emergence: The emergence score describes the ability of a seed to emergefrom the soil after planting. Each genotype is given a 1 to 9 scorebased on its percent of emergence. A score of 1 indicates an excellentrate and percent of emergence, an intermediate score of 5 indicates anaverage rating and a 9 score indicates a very poor rate and percent ofemergence.

Essentially all of the morphological and physiological characteristics:The characteristics of a plant are recovered that are otherwise presentwhen compared in the same environment, other than occasional varianttraits that might arise during backcrossing or direct introduction of atransgene.

FAME analysis: Fatty Acid Methyl Ester analysis is a method that allowsfor accurate quantification of the fatty acids that make up complexlipid classes.

Flower bud location: The location of the unopened flower buds relativeto the adjacent opened flowers is useful in distinguishing between thecanola species. The unopened buds are held above the most recentlyopened flowers in B. napus and they are positioned below the mostrecently opened flower buds in B. rapa.

Flowering date: This is measured by the number of days from planting tothe stage when 50% of the plants in a population have one or more openflowers. This varies from variety to variety.

Fusarium Wilt: Fusarium wilt, largely caused by Fusarium oxysporum, is adisease of canola that causes part or all of a plant to wilt, reducingyield by up to 30% or more on badly affected fields. For purposes ofthis application, resistance to Fusarium wilt is measured using ratingsof “R” (resistant), “MR” (medium resistant), “MS” (moderatelysusceptible) or “S” (susceptible).

Gene silencing: Gene silencing means the interruption or suppression ofthe expression of a gene at the level of transcription or translation.

Genotype: The genetic constitution of a cell or organism.

Glucosinolates: These are measured in micromoles (μm) of total alipathicglucosinolates per gram of air-dried oil-free meal. The level ofglucosinolates is somewhat influenced by the sulfur fertility of thesoil, but is also controlled by the genetic makeup of each variety andthus can be useful in characterizing varieties.

Growth habit: At the end of flowering, the angle relative to the groundsurface of the outermost fully expanded leaf petioles is a varietyspecific trait. This trait can range from erect (very upright along thestem) to prostrate (almost horizontal and parallel with the groundsurface).

Leaf attachment to the stem: This trait is especially useful fordistinguishing between the two canola species. The base of the leafblade of the upper stem leaves of B. rapa completely clasp the stemwhereas those of the B. napus only partially clasp the stem. Those ofthe mustard species do not clasp the stem at all.

Leaf blade color: The color of the leaf blades is variety specific andcan range from light to medium dark green to blue green.

Leaf development of lobes: The leaves on the upper portion of the stemcan show varying degrees of development of lobes which are disconnectedfrom one another along the petiole of the leaf. The degree of lobing isvariety specific and can range from absent (no lobes)/weak through verystrong (abundant lobes).

Leaf glaucosity: This refers to the waxiness of the leaves and ischaracteristic of specific varieties although environment can have someeffect on the degree of waxiness. This trait can range from absent (nowaxiness)/weak through very strong. The degree of waxiness can be bestdetermined by rubbing the leaf surface and noting the degree of waxpresent.

Leaf indentation of margin: The leaves on the upper portion of the stemcan also show varying degrees of serration along the leaf margins. Thedegree of serration or indentation of the leaf margins can vary fromabsent (smooth margin)/weak to strong (heavy saw-tooth like margin).

Leaf pubescence: The leaf pubescence is the degree of hairiness of theleaf surface and is especially useful for distinguishing between thecanola species. The two main classes of pubescence are glabrous(smooth/not hairy) and pubescent (hairy), which may be used todifferentiate between the B. napus and B. rapa species, respectively.

Leaf surface: The leaf surface can also be used to distinguish betweenvarieties. The surface can be smooth or rugose (lumpy) with varyingdegrees between the two extremes.

Linkage: A phenomenon wherein alleles on the same chromosome tend tosegregate together more often than expected by chance if theirtransmission was independent.

Linkage disequilibrium: A phenomenon wherein alleles tend to remaintogether in linkage groups when segregating from parents to offspring,with a greater frequency than expected from their individualfrequencies.

Locus: A locus confers one or more traits such as, for example, malesterility, herbicide tolerance, insect resistance, disease resistance,modified fatty acid metabolism, modified phytic acid metabolism,modified carbohydrate metabolism and modified protein metabolism. Thetrait may be, for example, conferred by a naturally occurring geneintroduced into the genome of the variety by backcrossing, a natural orinduced mutation, or a transgene introduced through genetictransformation techniques. A locus may comprise one or more allelesintegrated at a single chromosomal location.

Lodging resistance: Lodging is rated on a scale of 1 to 5. A score of 1indicates that the plants are erect. A score of 5 indicates that theplants are lying on the ground

Maturity: The maturity of a variety is measured as the number of daysbetween planting and physiological maturity. This is useful trait indistinguishing varieties relative to one another.

Moisture: The average percentage moisture in the seeds of the variety.

Oil content: This is measured as percent of the whole dried seed and ischaracteristic of different varieties. It can be determined usingvarious analytical techniques such as NMR, NIR, and Soxhlet extraction.

Oil or Oil Percent: Seed oil content is measured and reported on apercentage basis.

Percent linolenic acid: Percent oil of the seed that is linolenic acid.

Percent oleic acid (OLE): Percent oil of the seed that is oleic acid.

Percentage of total fatty acids: This is determined by extracting asample of oil from seed, producing the methyl esters of fatty acidspresent in that oil sample and analyzing the proportions of the variousfatty acids in the sample using gas chromatography. The fatty acidcomposition can also be a distinguishing characteristic of a variety.

Petal color: The petal color on the first day a flower opens can be adistinguishing characteristic for a variety. It can be white, varyingshades of yellow or orange.

Plant: As used herein, the term “plant” includes reference to animmature or mature whole plant, including a plant from which seed orgrain or anthers have been removed. Seed or embryo that will produce theplant is also considered to be the plant.

Plant height: This is the height of the plant at the end of flowering ifthe floral branches are extended upright (i.e., not lodged). This variesfrom variety to variety and although it can be influenced byenvironment, relative comparisons between varieties grown side by sideare useful for variety identification.

Plant parts: As used herein, the term “plant parts” (or a canola plant,or a part thereof) includes protoplasts, spheroplasts, leaves, stems,roots, root tips, anthers, pistils, seed, grain, embryo, pollen, ovules,cotyledon, hypocotyl, pod, flower, shoot, tissue, petiole, cells,meristematic cells and the like.

Protein content: This is measured as percent of whole dried seed and ischaracteristic of different varieties. This can be determined usingvarious analytical techniques such as NIR and Kjeldahl.

Quantitative trait loci (QTL): A genetic loci that control to somedegree numerically representable traits that are usually continuouslydistributed.

Regeneration: The development of a plant from tissue culture.

Resistance to lodging: This measures the ability of a variety to standup in the field under high yield conditions and severe environmentalfactors. A variety can have good (remains upright), fair, or poor (fallsover) resistance to lodging. The degree of resistance to lodging is notexpressed under all conditions but is most meaningful when there is somedegree of lodging in a field trial.

Seed coat color: The color of the seed coat can be variety specific andcan range from black through brown through yellow. Color can also bemixed for some varieties.

Seed coat mucilage: This is useful for differentiating between the twospecies of canola with B. rapa varieties have mucilage present in theirseed coats; whereas, B. napus varieties do not have this present. It isdetected by imbibing seeds with water and monitoring the mucilage thatis exuded by the seed.

Seedling growth habit: The rosette consists of the first 2-8 true leavesand a variety can be characterized as having a strong rosette (closelypacked leaves) or a weak rosette (loosely arranged leaves).

Silique (pod) habit: A trait that is variety specific and is a measureof the orientation of the pods along the racemes (flowering stems). Thistrait can range from erect (pods angled close to racemes) throughhorizontal (pods perpendicular to racemes) through arching (pods showdistinct arching habit).

Silique (pod) length of beak: The beak is the segment at the end of thepod which does not contain seed (it is a remnant of the stigma and stylefor the flower). The length of the beak can be variety specific and canrange from short through medium through long.

Silique (pod) length of pedicel: The pedicel is the stem that attachesthe pod to the raceme of flowering shoot. The length of the pedicel canbe variety specific and can vary from short through medium through long.

Silique (pod) length: This is the length of the fully developed pods andcan range from short to medium to long. It is best used by makingcomparisons relative to reference varieties.

Silique (pod) type: This is typically a bilateral single pod for bothspecies of canola and is not really useful for variety identificationwithin these species.

Silique (pod) width: This is the width of the fully developed pods andcan range from narrow to medium to wide. It is best used by makingcomparisons relative to reference varieties.

Single locus converted (conversion) plant: Plants that are developed bya plant breeding technique called backcrossing or by genome editing of alocus, wherein essentially all of the morphological and physiologicalcharacteristics of a variety are recovered in addition to the singlelocus transferred into the variety via the backcrossing technique or bygenetic transformation. It is understood that once introduced into anyplant genome, a locus that is transgenic in origin (transgene), can beintroduced by backcrossing as with any other locus.

Stem intensity of anthocyanin coloration: The stems and other organs ofcanola plants can have varying degrees of purple coloration which is dueto the presence of anthocyanin (purple) pigments. The degree ofcoloration is somewhat subject to growing conditions, but varietiestypically show varying degrees of coloration ranging from: absent (nopurple)/very weak to very strong (deep purple coloration).

Tissue culture: A composition comprising isolated cells of the same or adifferent type or a collection of such cells organized into parts of aplant.

Total saturated (TOTSAT): Total percent oil of the seed of the saturatedfats in the oil including C12:0, C14:0, C16:0, C18:0, C20:0, C22:0 andC24.0.

Transgene: A genetic locus comprising a sequence which has beenintroduced into the genome of a canola plant by transformation orsite-specific recombination.

DETAILED DESCRIPTION OF THE INVENTION

Spring canola variety SCV528587 was developed from the initial cross(CK6686:0002.0001.0008. @0.0003)/(65037*3/(WINNER/MENDEL:0081%0001.@!):103065>101270>2387.120149.@0.0004).

Generation Year Description Cross 2009 The cross was made near Winnipeg,Manitoba. F1 2011 F1 plant grown in Winnipeg growthroom as microsporedonor, planted January 2011. DH0 2011 DH0 plants selfed in Winnipeggrowthroom July 2011 to produce DH1 seed. DH1 2012 DH1 plants selfed inChile January 2012 to produce DH2 seed. DH2 2012 DH2 plants selfed inChile January 2012 to produce DH3 seed. DH3 2014 DH3 plants selfed inWinnipeg growthroom November 2014 to produce DH4 seed. DH4 2017 DH4plants bulk pollinated in Chile January 2017 to produce DH5 breederseed. Variety SCV528587 was selected.

Canola line SCV528587 is stable and uniform and no off-type plants havebeen exhibited in evaluation. The line has shown uniformity andstability, as described in the following variety descriptioninformation. It has been selfed a sufficient number of generations withcareful attention to uniformity of plant type. The line has beenincreased with continued observation for uniformity.

Canola variety SCV528587 has the following morphological and othercharacteristics:

TABLE 1 Phenotypic Description of Canola Variety SCV528587 TraitPhenotype Classification: Species Brassica napus L. Season Type Springhabit Type of pollination control Cytoplasmic male sterility: INRA Oguratype Characteristics of Plants Before Flowering: Cotyledon width MediumSeedling growth habit (leaf rosette) Medium/small Stem anthocyaninintensity Nearly Absent Leaf type Intermediate to petiolate - lyrateLeaf shape Narrow elliptic Leaf color Medium green Leaf waxiness MediumLeaf margin indentation Strong (deep) Leaf attachment to the stemPartial clasping Characteristics of Plants After Flowering: Time toflowering 48 Plant height at maturity Short-Medium Plant growth habitIntermediate Flower bud location Buds above most recently opened flowersPetal color Dark yellow Anther arrangement Introrse Pod lengthMedium/short Pod angle Semi erect Pod beak length Medium Pedicel lengthMedium Time to maturity 94 Seed Characteristics Seed coat colorBlack/brown Disease & Pest Reactions: Blackleg Resistant Club RootResistant Fusarium wilt Resistant Herbicide Reactions: GlyphosateSusceptible

Public or Commercial Designations Used for the Original Parent Lines:

Canola variety 65037 is also known as SCV378221. WINNER and MENDEL arecommercial winter oilseed rape varieties.

Related Art:

No patent applications have been filed or patents issued in which anysiblings of canola variety SCV528587 are claimed.

This invention is also directed to methods for producing a canola plantby crossing a first parent canola plant with a second parent canolaplant, wherein the first or second canola plant is the canola plant fromthe variety SCV528587. Further, both first and second parent canolaplants may be from the variety SCV528587. Therefore, any methods usingthe variety SCV528587 are part of this invention: selfing, backcrosses,hybrid breeding, and crosses to populations. Any plants produced usingvariety SCV528587 as a parent are within the scope of this invention.

Additional methods include, but are not limited to, expression vectorsintroduced into plant tissues using a direct gene transfer method suchas microprojectile-mediated delivery, DNA injection, electroporation,and the like. More preferably, expression vectors may be introduced intoplant tissues by using either microprojectile-mediated delivery with aballistic device or by using Agrobacterium-mediated transformation.Transformant plants obtained with the protoplasm of the invention areintended to be within the scope of this invention.

The advent of new molecular biological techniques has allowed theisolation and characterization of genetic elements with specificfunctions, such as encoding specific protein products. Scientists in thefield of plant biology developed a strong interest in engineering thegenome of plants to contain and express foreign genetic elements, oradditional, or modified versions of native or endogenous geneticelements in order to alter the traits of a plant in a specific manner.Any DNA sequences, whether from a different species or from the samespecies which are inserted into the genome using transformation, arereferred to herein collectively as “transgenes.” In some embodiments ofthe invention, a transgenic variant of SCV528587 may contain at leastone transgene but could contain at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10and/or no more than 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, or 2.Over the last fifteen to twenty years, several methods for producingtransgenic plants have been developed, and the present invention alsorelates to transgenic variants of the claimed canola variety SCV528587.

One embodiment of the invention is a process for producing canolavariety SCV528587 further comprising a trait, said process comprisingtransforming a canola plant of variety SCV528587 with a transgene thatconfers a trait. Another embodiment is the product produced by thisprocess. In one embodiment the trait may be one or more of herbicideresistance, insect resistance, disease resistance, modified seed yield,modified oil percent, modified protein percent, modified lodgingresistance or modified fatty acid or carbohydrate metabolism. Thespecific gene may be any known in the art or listed herein, including; apolynucleotide conferring resistance to imidazolinone, sulfonylurea,glyphosate, glufosinate, triazine, hydroxyphenylpyruvate dioxygenaseinhibitor, protoporphyrinogen oxidase inhibitor and benzonitrile; apolynucleotide encoding a Bacillus thuringiensis polypeptide, apolynucleotide encoding phytase, FAD-2, FAD-3, galactinol synthase or araffinose synthetic enzyme; or a polynucleotide conferring resistance toblackleg, white rust or other common canola diseases.

Numerous methods for plant transformation have been developed, includingbiological and physical plant transformation protocols, all of which maybe used with this invention. In addition, expression vectors and invitro culture methods for plant cell or tissue transformation andregeneration of plants are available and may be used in conjunction withthe invention.

In an embodiment, a genetic trait which has been engineered into thegenome of a particular canola plant may be moved into the genome ofanother variety using traditional breeding techniques that are wellknown in the plant breeding arts. For example, a backcrossing approachmay be used to move a transgene from a transformed canola variety intoan already developed canola variety, and the resulting backcrossconversion plant would then comprise the transgene(s).

In embodiments, various genetic elements can be introduced into theplant genome using transformation. These elements include any known inthe art, specifically including, but not limited to genes, codingsequences, inducible, constitutive, and tissue specific promoters,enhancing sequences, and signal and targeting sequences.

Plant transformation involves the construction of an expression vectorwhich will function in plant cells. Such a vector comprises DNAcomprising a gene under control of or operatively linked to a regulatoryelement (for example, a promoter). The expression vector may contain oneor more of such operably linked gene/regulatory element combinations.The vector(s) may be in the form of a plasmid, and can be used alone orin combination with other plasmids, to provide transformed canolaplants, using transformation methods as described below to incorporatetransgenes into the genetic material of the canola plant(s).

Expression Vectors for Canola Transformation: Marker Genes

Expression vectors include at least one genetic marker operably linkedto a regulatory element (a promoter, for example) that allowstransformed cells containing the marker to be either recovered bynegative selection, i.e., inhibiting growth of cells that do not containthe selectable marker gene, or by positive selection, i.e., screeningfor the product encoded by the genetic marker. Many commonly usedselectable marker genes for plant transformation are well known in thetransformation arts, and include, for example, genes that code forenzymes that metabolically detoxify a selective chemical agent which maybe an antibiotic or an herbicide, or genes that encode an altered targetwhich is insensitive to the inhibitor. A few positive selection methodsare also known in the art.

One commonly used selectable marker gene for plant transformation is theneomycin phosphotransferase II (nptII) gene which, when under thecontrol of plant regulatory signals, confers resistance to kanamycin.Another commonly used selectable marker gene is the hygromycinphosphotransferase gene which confers resistance to the antibiotichygromycin.

Additional selectable marker genes of bacterial origin that conferresistance to antibiotics include gentamycin acetyl transferase,streptomycin phosphotransferase, aminoglycoside-3′-adenyl transferaseand the bleomycin resistance determinant. Other selectable marker genesconfer resistance to herbicides such as glyphosate, glufosinate orbromoxynil. Selectable marker genes for plant transformation not ofbacterial origin include, for example, mouse dihydrofolate reductase,plant 5-enolpyruvylshikimate-3-phosphate synthase and plant acetolactatesynthase.

Another class of marker genes for plant transformation requiresscreening of presumptively transformed plant cells rather than directgenetic selection of transformed cells for resistance to a toxicsubstance such as an antibiotic. These genes are particularly useful toquantify or visualize the spatial pattern of expression of a gene inspecific tissues and are frequently referred to as reporter genesbecause they can be fused to a gene or gene regulatory sequence for theinvestigation of gene expression. Commonly used genes for screeningpresumptively transformed cells include β-glucuronidase (GUS),β-galactosidase, luciferase and chloramphenicol acetyltransferase. Anyof the above, or other marker genes, may be utilized in the presentinvention.

In vivo methods for visualizing GUS activity that do not requiredestruction of plant tissue are available and can be used in embodimentsof the invention. Additionally, Green Fluorescent Protein (GFP) can beutilized as a marker for gene expression in prokaryotic and eukaryoticcells. GFP and mutants of GFP may be used as screenable markers.

Expression Vectors for Canola Transformation: Promoters

Genes included in expression vectors must be driven by a nucleotidesequence comprising a regulatory element, for example, a promoter.Several types of promoters are well known in the transformation arts, asare other regulatory elements that can be used alone or in combinationwith promoters.

As used herein, “promoter” includes reference to a region of DNAupstream from the start of transcription and involved in recognition andbinding of RNA polymerase and other proteins to initiate transcription.A “plant promoter” is a promoter capable of initiating transcription inplant cells. Examples of promoters under developmental control includepromoters that preferentially initiate transcription in certain tissues,such as leaves, roots, seeds, fibers, xylem vessels, tracheids, orsclerenchyma. Such promoters are referred to as “tissue-preferred”.Promoters which initiate transcription only in certain tissues arereferred to as “tissue-specific”. A “cell type” specific promoterprimarily drives expression in certain cell types in one or more organs,for example, vascular cells in roots or leaves. An “inducible” promoteris a promoter which is under environmental control. Examples ofenvironmental conditions that may affect transcription by induciblepromoters include anaerobic conditions or the presence of light.Tissue-specific, tissue-preferred, cell type specific, and induciblepromoters constitute the class of “non-constitutive” promoters. A“constitutive” promoter is a promoter which is active under mostenvironmental conditions.

Inducible Promoters—An inducible promoter is operably linked to a genefor expression in canola. Optionally, the inducible promoter is operablylinked to a nucleotide sequence encoding a signal sequence which isoperably linked to a gene for expression in canola. With an induciblepromoter the rate of transcription increases in response to an inducingagent.

Any inducible promoter can be used in the instant invention. Exemplaryinducible promoters include, but are not limited to, those from the ACEIsystem which respond to copper, the In2 gene from maize which respondsto benzenesulfonamide herbicide safeners, or the Tet repressor fromTn10. A particularly preferred inducible promoter is a promoter thatresponds to an inducing agent to which plants do not normally respond.An exemplary inducible promoter is the inducible promoter from a steroidhormone gene, the transcriptional activity of which is induced by aglucocorticosteroid hormone.

Constitutive Promoters—A constitutive promoter is operably linked to agene for expression in canola or the constitutive promoter is operablylinked to a nucleotide sequence encoding a signal sequence which isoperably linked to a gene for expression in canola.

Many different constitutive promoters can be utilized in the instantinvention. Exemplary constitutive promoters include, but are not limitedto, the promoters from plant viruses such as the 35S promoter from CaMVand the promoters from such genes as rice actin, ubiquitin, pEMU, MAS,and maize H3 histone. The ALS promoter, Xba1/Nco1 fragment 5′ to theBrassica napus ALS3 structural gene (or a nucleotide sequence similarityto said Xba1/Nco1 fragment) could also be utilized herein.

Tissue-specific or Tissue-preferred Promoters—A tissue-specific promoteris operably linked to a gene for expression in canola. Optionally, thetissue-specific promoter is operably linked to a nucleotide sequenceencoding a signal sequence which is operably linked to a gene forexpression in canola. Plants transformed with a gene of interestoperably linked to a tissue-specific promoter produce the proteinproduct of the transgene exclusively, or preferentially, in a specifictissue.

Any tissue-specific or tissue-preferred promoter can be utilized in theinstant invention. Exemplary tissue-specific or tissue-preferredpromoters include, but are not limited to, a root-preferred promotersuch as that from the phaseolin gene, a leaf-specific and light-inducedpromoter such as that from cab or rubisco, an anther-specific promotersuch as that from LAT52, a pollen-specific promoter such as that fromZm13, or a microspore-preferred promoter such as that from apg.

Signal Sequences for Targeting Proteins to Subcellular Compartments

Transport of protein produced by transgenes to a subcellular compartmentsuch as the chloroplast, vacuole, peroxisome, glyoxysome, cell wall ormitochondrion or for secretion into the apoplast, is accomplished bymeans of operably linking the nucleotide sequence encoding a signalsequence to the 5′ and/or 3′ region of a gene encoding the protein ofinterest. Targeting sequences at the 5′ and/or 3′ end of the structuralgene may determine, during protein synthesis and processing, where theencoded protein is ultimately compartmentalized. The presence of asignal sequence directs a polypeptide to either an intracellularorganelle or subcellular compartment or for secretion to the apoplast.Many signal sequences are known in the art and can be utilized in thepresent invention.

Foreign Protein Genes and Agronomic Genes

With transgenic plants according to the present invention, a foreignprotein can be produced in commercial quantities. Thus, techniques forthe selection and propagation of transformed plants, which are wellunderstood in the art, are within the scope of the invention. In anembodiment, a foreign protein then can be extracted from a tissue ofinterest or from the total biomass by known methods.

According to a preferred embodiment, the transgenic plant provided forcommercial production of foreign protein is a canola plant. In anotherpreferred embodiment, the biomass of interest is seed. For therelatively small number of transgenic plants that show higher levels ofexpression, a genetic map can be generated, primarily via conventionalRFLP, PCR and SSR analysis, which identifies the approximate chromosomallocation of the integrated DNA molecule. Map information concerningchromosomal location is useful for proprietary protection of a subjecttransgenic plant. If unauthorized propagation is undertaken and crossesare made with other germplasm, the map of the integration region can becompared to similar maps for suspect plants, to determine if the latterhave a common parentage with the subject plant. Map comparisons wouldinvolve hybridizations, RFLP, PCR, SSR and sequencing, all of which areconventional techniques. SNPs may also be used alone or in combinationwith other techniques.

Likewise, by means of the present invention, plants can be geneticallyengineered to express various phenotypes of agronomic interest. Throughthe transformation of canola, the expression of genes can be altered toenhance disease resistance, insect resistance, herbicide resistance,agronomic, grain quality and other traits. Transformation can also beused to insert DNA sequences which control, or help control,male-sterility. DNA sequences native to canola, as well as non-nativeDNA sequences, can be transformed into canola and used to alter levelsof native or non-native proteins. Various promoters, targetingsequences, enhancing sequences, and other DNA sequences can be insertedinto the genome for the purpose of altering the expression of proteins.Reduction of the activity of specific genes (also known as genesilencing, or gene suppression) is desirable for several aspects ofgenetic engineering in plants.

Many techniques for gene silencing are well known to one of skill in theart, including but not limited to knock-outs (such as by insertion of atransposable element such as Mu or other genetic elements such as a FRT,Lox or other site specific integration site), antisense technology,co-suppression, RNA interference, virus-induced gene silencing,target-RNA-specific ribozymes, hairpin structures, MicroRNA, ribozymes,oligonucleotide-mediated targeted modification, Zn-finger targetedmolecules, and other methods or combinations of the above methods knownto those of skill in the art.

Likewise, by means of the present invention, agronomic genes can beexpressed in transformed plants. More particularly, plants can begenetically engineered to express various phenotypes of agronomicinterest. Exemplary genes implicated in this regard include, but are notlimited to, those categorized below:

A. Genes that Confer Resistance to Pests or Disease and that Encode:

Plant defenses are often activated by specific interaction between theproduct of a disease resistance gene (R) in the plant and the product ofa corresponding avirulence (Avr) gene in the pathogen. A plant line canbe transformed with cloned resistance genes to engineer plants that areresistant to specific pathogen strains.

A viral-invasive protein or a complex toxin derived therefrom may alsobe used for viral disease resistance. For example, the accumulation ofviral coat proteins in transformed plant cells imparts resistance toviral infection and/or disease development effected by the virus fromwhich the coat protein gene is derived, as well as by and relatedviruses. Coat protein-mediated resistance has been conferred upontransformed plants against alfalfa mosaic virus, cucumber mosaic virus,tobacco streak virus, potato virus X, potato virus Y, tobacco etchvirus, tobacco rattle virus, and tobacco mosaic virus.

A virus-specific antibody may also be used. See, for example,Tavladoraki et al. (Nature, 366:469-472, 1993), who show that transgenicplants expressing recombinant antibody genes are protected from virusattack. Virus resistance has also been described in, for example, U.S.Pat. Nos. 6,617,496; 6,608,241; 6,015,940; 6,013,864; 5,850,023 and5,304,730. Additional means of inducing whole-plant resistance to apathogen include modulation of the systemic acquired resistance (SAR) orpathogenesis related (PR) genes, for example genes homologous to theArabidopsis thaliana NIM1/NPR1/SAI1, and/or by increasing salicylic acidproduction (Ryals et al., Plant Cell, 8:1809-1819, 1996).

A gene conferring resistance to fungal pathogens, such as oxalateoxidase or oxalate decarboxylase.

A Bacillus thuringiensis protein, a derivative thereof, or a syntheticpolypeptide modeled thereon, for example, a Bt δ-endotoxin gene.

A lectin.

A vitamin-binding protein such as avidin or a homolog.

An enzyme inhibitor, for example, a protease or proteinase inhibitor oran amylase inhibitor.

An insect-specific hormone or pheromone such as an ecdysteroid orjuvenile hormone, a variant thereof, a mimetic based thereon, or anantagonist or agonist thereof.

An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest.

An insect-specific venom produced in nature by a snake, a wasp, etc.

An enzyme responsible for a hyperaccumulation of a monoterpene, asesquiterpene, a steroid, hydroxamic acid, a phenylpropanoid derivativeor another non-protein molecule with insecticidal activity.

An enzyme involved in the modification, including the post-translationalmodification, of a biologically active molecule; for example, aglycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme, a nuclease,a cyclase, a transaminase, an esterase, a hydrolase, a phosphatase, akinase, a phosphorylase, a polymerase, an elastase, a chitinase and aglucanase, whether natural or synthetic.

A molecule that stimulates signal transduction.

A hydrophobic moment peptide.

A membrane permease, a channel former or a channel blocker.

A viral-invasive protein or a complex toxin derived therefrom. Forexample, the accumulation of viral coat proteins in transformed plantcells imparts resistance to viral infection and/or disease developmenteffected by the virus from which the coat protein gene is derived, aswell as by related viruses. Coat protein-mediated resistance has beenconferred upon transformed plants against alfalfa mosaic virus, cucumbermosaic virus, tobacco streak virus, potato virus X, potato virus Y,tobacco etch virus, tobacco rattle virus and tobacco mosaic virus.

An insect-specific antibody or an immunotoxin derived therefrom. Anantibody targeted to a critical metabolic function in the insect gutwould inactivate an affected enzyme, killing the insect.

A virus-specific antibody.

A developmental-arrestive protein produced in nature by a pathogen or aparasite. Thus, fungal endo-α-1, 4-D-polygalacturonases facilitatefungal colonization and plant nutrient release by solubilizing plantcell wall homo-α-1, 4-D-galacturonase.

A developmental-arrestive protein produced in nature by a plant.

Genes involved in the Systemic Acquired Resistance (SAR) Response and/orthe pathogenesis-related genes.

Antifungal genes.

Detoxification genes, such as for fumonisin, beauvericin, moniliforminand zearalenone and their structurally related derivatives.

Cystatin and cysteine proteinase inhibitors.

Defensin genes.

Genes that confer resistance to Phytophthora root rot, such as theBrassica equivalents of the Rps 1, Rps 1-a, Rps 1-b, Rps 1-c, Rps 1-d,Rps 1-e, Rps 1-k, Rps 2, Rps 3-a, Rps 3-b, Rps 3-c, Rps 4, Rps 5, Rps 6,Rps 7 and other Rps genes.

B. Genes that Confer Resistance to an Herbicide, for Example:

Numerous herbicide resistance genes are known and may be employed withthe invention. A non-limiting example is a gene conferring resistance toa herbicide that inhibits the growing point or meristem such asimidazolinone or sulfonylurea herbicides. As imidazolinone andsulfonylurea herbicides are acetolactate synthase (ALS)-inhibitingherbicides that prevent the formation of branched chain amino acids,exemplary genes in this category code for ALS and AHAS enzymes asdescribed, for example, by Lee et al., EMBO J., 7:1241, 1988; Gleen etal., Plant Molec. Biology, 18:1185, 1992; and Miki et al., Theor. Appl.Genet., 80:449, 1990. As a non-limiting example, a gene may be employedto confer resistance to the exemplary sulfonylurea herbicidenicosulfuron.

Resistance genes for glyphosate (resistance conferred by mutant5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) and aroA genes,respectively) and other phosphono compounds such as glufosinate(phosphinothricin acetyltransferase (PAT) and Streptomyces hygroscopicusPAT bar genes) may also be used. See, for example, U.S. Pat. No.4,940,835 to Shah et al., which discloses the nucleotide sequence of aform of EPSPS that can confer glyphosate resistance. Non-limitingexamples of EPSPS transformation events conferring glyphosate resistanceare provided by U.S. Pat. Nos. 6,040,497 and 7,632,985. The MON89788event disclosed in U.S. Pat. No. 7,632,985 in particular is beneficialin conferring glyphosate tolerance in combination with an increase inaverage yield relative to prior events.

A DNA molecule encoding a mutant aroA gene can be obtained under ATCCAccession No. 39256, and the nucleotide sequence of the mutant gene isdisclosed in U.S. Pat. No. 4,769,061 to Comai. A hygromycin Bphosphotransferase gene from E. coli that confers resistance toglyphosate in tobacco callus and plants is described in Penaloza-Vazquezet al., Plant Cell Reports, 14:482, 1995. European Patent ApplicationPublication No. EP0333033 to Kumada et al., and U.S. Pat. No. 4,975,374to Goodman et al., disclose nucleotide sequences of glutamine synthetasegenes that confer resistance to herbicides such as L-phosphinothricin.The nucleotide sequence of a phosphinothricin acetyltransferase gene isprovided in European Patent Application Publication No. EP0242246 toLeemans et al. DeGreef et al. (Biotechnology, 7:61, 1989) describe theproduction of transgenic plants that express chimeric bar genes codingfor phosphinothricin acetyl transferase activity. Exemplary genesconferring resistance to a phenoxy class herbicide haloxyfop and acyclohexanedione class herbicide sethoxydim are the Acct-S1, Acct-S2 andAcct-S3 genes described by Marshall et al., (Theor. Appl. Genet.,83:435, 1992). As a non-limiting example, a gene may confer resistanceto other exemplary phenoxy class herbicides that include, but are notlimited to, quizalofop-p-ethyl and 2,4-dichlorophenoxyacetic acid(2,4-D).

Genes are also known that confer resistance to herbicides that inhibitphotosynthesis, such as, for example triazine herbicides (psbA and gs+genes) and benzonitrile herbicides (nitrilase gene). Przibila et al.(Plant Cell, 3:169, 1991) describe the transformation of Chlamydomonaswith plasmids encoding mutant psbA genes. Nucleotide sequences fornitrilase genes are disclosed in U.S. Pat. No. 4,810,648 to Stalker, andDNA molecules containing these genes are available under ATCC AccessionNos. 53435, 67441, and 67442. Cloning and expression of DNA coding for aglutathione S-transferase is described by Hayes et al. (Biochem. J.,285:173, 1992). 4-hydroxyphenylpyruvate dioxygenase (HPPD) is a targetof the HPPD-inhibiting herbicides, which deplete plant plastoquinone andvitamin E pools. Rippert et al. (Plant Physiol., 134:92, 2004) describesan HPPD-inhibitor resistant tobacco plant that was transformed with ayeast-derived prephenate dehydrogenase (PDH) gene. Protoporphyrinogenoxidase (PPO) is the target of the PPO-inhibitor class of herbicides; aPPO-inhibitor resistant PPO gene was recently identified in Amaranthustuberculatus (Patzoldt et al., PNAS, 103(33):12329, 2006). The herbicidemethyl viologen inhibits CO₂ assimilation. Foyer et al. (Plant Physiol.,109:1047, 1995) describe a plant overexpressing glutathione reductase(GR) that is resistant to methyl viologen treatment.

Siminszky (Phytochemistry Reviews, 5:445, 2006) describes plantcytochrome P450-mediated detoxification of multiple, chemicallyunrelated classes of herbicides. Modified bacterial genes have beensuccessfully demonstrated to confer resistance to atrazine, a herbicidethat binds to the plastoquinone-binding membrane protein Q_(B) inphotosystem II to inhibit electron transport. See, for example, studiesby Cheung et al. (PNAS, 85:391, 1988), describing tobacco plantsexpressing the chloroplast psbA gene from an atrazine-resistant biotypeof Amaranthus hybridus fused to the regulatory sequences of a nucleargene, and Wang et al. (Plant Biotech. J., 3:475, 2005), describingtransgenic alfalfa, Arabidopsis, and tobacco plants expressing the atzAgene from Pseudomonas sp. that were able to detoxify atrazine.

Bayley et al. (Theor. Appl. Genet., 83:645, 1992) describe the creationof 2,4-D-resistant transgenic tobacco and cotton plants using the 2,4-Dmonooxygenase gene OA from Alcaligenes eutrophus plasmid pJP5. U.S.Patent Application Publication No. 20030135879 describes the isolationof a gene for dicamba monooxygenase (DMO) from Psueodmonas maltophiliathat is involved in the conversion of dicamba to a non-toxic3,6-dichlorosalicylic acid and thus may be used for producing plantstolerant to this herbicide.

Other examples of herbicide resistance have been described, forinstance, in U.S. Pat. Nos. 6,803,501; 6,448,476; 6,248,876; 6,225,114;6,107,549; 5,866,775; 5,804,425; 5,633,435; 5,463,175.

C. Genes that Confer or Contribute to a Value-Added Trait, Such as:

Modified fatty acid metabolism, for example, by transforming a plantwith an antisense gene of stearyl-ACP desaturase to increase stearicacid content of the plant. Decreased phytate content. Introduction of aphytase-encoding gene, such as Aspergillus niger phytase gene, mayenhance breakdown of phytate, adding more free phosphate to thetransformed plant. Alternatively, a gene could be introduced thatreduces phytate content. In maize for example, this could beaccomplished by cloning and then reintroducing DNA associated with thesingle allele which is responsible for maize mutants characterized bylow levels of phytic acid.

Modified carbohydrate composition effected, for example, by transformingplants with a gene coding for an enzyme that alters the branchingpattern of starch, or, a gene altering thioredoxin such as NTR and/orTRX and/or a gamma zein knock out or mutant such as cs27 or TUSC27 oren27. Any known fatty acid modification genes may also be used to affectstarch content and/or composition through the interrelationship of thestarch and oil pathways.

Elevated oleic acid via FAD-2 gene modification and/or decreasedlinolenic acid via FAD-3 gene modification.

Altering conjugated linolenic or linoleic acid content. Altering LEC1,AGP, Dek1, Superal1, mi1ps, various Ipa genes such as Ipa1, Ipa3, hpt orhggt.

Altered antioxidant content or composition, such as alteration oftocopherol or tocotrienols. In an embodiment, antioxidant levels may bemanipulated through alteration of a phytl prenyl transferase (ppt) orthrough alteration of a homogentisate geranyl geranyl transferase(hggt).

Altered essential seed amino acids.

D. Genes that Control Male Sterility

There are several methods of conferring genetic male sterility availableand within the scope of the invention. As one example, nuclear malesterility may be accomplished by identifying a gene which is critical tomale fertility, silencing this native gene which is critical to malefertility, removing the native promoter from the essential malefertility gene and replacing it with an inducible promoter, insertingthis genetically engineered gene back into the plant, and thus creatinga plant that is male sterile because the inducible promoter is not “on,”resulting in the male fertility gene not being transcribed. Fertility isrestored by inducing, or turning “on”, the promoter, which in turnallows the gene that confers male fertility to be transcribed. Otherpossible examples include the introduction of a deacetylase gene underthe control of a tapetum-specific promoter and with the application ofthe chemical N-Ac-PPT, the introduction of various stamen-specificpromoters, or the introduction of the barnase and the barstar genes.

E. Genes that Create a Site for Site Specific DNA Integration.

This may include the introduction of FRT sites that may be used in theFLP/FRT system and/or Lox sites that may be used in the Cre/Loxp system.Other systems that may be used include the Gin recombinase of phage Mu,the Pin recombinase of E. coli, and the R/RS system of the pSR1 plasmid.

F. Genes that Affect Abiotic Stress Resistance (Including but notLimited to Flowering, Pod and Seed Development, Enhancement of NitrogenUtilization Efficiency, Altered Nitrogen Responsiveness, DroughtResistance or Tolerance, Cold Resistance or Tolerance, and SaltResistance or Tolerance) and Increased Yield Under Stress.

Abiotic stress includes dehydration or other osmotic stress, salinity,high or low light intensity, high or low temperatures, submergence,exposure to heavy metals, and oxidative stress.Delta-pyrroline-5-carboxylate synthetase (P5CS) from mothbean has beenused to provide protection against general osmotic stress.Mannitol-1-phosphate dehydrogenase (mt1D) from E. coli has been used toprovide protection against drought and salinity. Choline oxidase (codAfrom Arthrobactor globiformis) can protect against cold and salt. E.coli choline dehydrogenase (betA) provides protection against salt.Additional protection from cold can be provided by omega-3-fatty aciddesaturase (fad7) from Arabidopsis thaliana. Trehalose-6-phosphatesynthase and levan sucrase (SacB) from yeast and Bacillus subtilis,respectively, can provide protection against drought (summarized fromAnnex II Genetic Engineering for Abiotic Stress Tolerance in Plants,Consultative Group On International Agricultural Research TechnicalAdvisory Committee). Overexpression of superoxide dismutase can be usedto protect against superoxides, see U.S. Pat. No. 5,538,878. Genes andtranscription factors that affect plant growth and agronomic traits suchas yield, flowering, plant growth and/or plant structure, can beintroduced or introgressed into plants.

Methods for Canola Transformation

Numerous methods for plant transformation have been developed, includingbiological and physical plant transformation protocols. In addition,expression vectors and in vitro culture methods for plant cell or tissuetransformation and regeneration of plants are available.

Agrobacterium-mediated Transformation—One method for introducing anexpression vector into plants is based on the natural transformationsystem of Agrobacterium. A. tumefaciens and A. rhizogenes are plantpathogenic soil bacteria which genetically transform plant cells. The Tiand Ri plasmids of A. tumefaciens and A. rhizogenes, respectively, carrygenes responsible for genetic transformation of the plant. Agrobacteriumvector systems and methods for Agrobacterium-mediated gene transfer canbe used in the present invention.

Direct Gene Transfer—Several methods of plant transformation,collectively referred to as direct gene transfer, have been developed asan alternative to Agrobacterium-mediated transformation. A generallyapplicable method of plant transformation is microprojectile-mediatedtransformation wherein DNA is carried on the surface of microprojectilesmeasuring 1 to 4 μm. The expression vector is introduced into planttissues with a ballistic device that accelerates the microprojectiles tospeeds of 300 to 600 m/s which is sufficient to penetrate plant cellwalls and membranes. Another method for physical delivery of DNA toplants is sonication of target cells, which may be used herein.Alternatively, liposome and spheroplast fusion may be used to introduceexpression vectors into plants. Direct uptake of DNA into protoplastsusing CaCl₂ precipitation, polyvinyl alcohol or poly-L-ornithine mayalso be useful. Electroporation of protoplasts and whole cells andtissues may also be utilized.

Included among various plant transformation techniques are methodspermitting the site-specific modification of a plant genome. Thesemodifications can include, but are not limited to, site-specificmutations, deletions, insertions, and replacements of nucleotides. Thesemodifications can be made anywhere within the genome of a plant, forexample, in genomic elements, including, among others, coding sequences,regulatory elements, and non-coding DNA sequences. Any number of suchmodifications can be made and that number of modifications may be madein any order or combination, for example, simultaneously all together orone after another. Such methods may be used to modify a particular traitconferred by a locus. The techniques for making such modifications bygenome editing are well known in the art and include, for example, useof CRISPR-Cas systems, zinc-finger nucleases (ZFNs), and transcriptionactivator-like effector nucleases (TALENs), among others.

Following transformation of canola target tissues, expression of theabove-described selectable marker genes allows for preferentialselection of transformed cells, tissues and/or plants, usingregeneration and selection methods well known in the art.

The foregoing methods for transformation would typically be used forproducing a transgenic variety. The transgenic variety could then becrossed with another (non-transformed or transformed) variety in orderto produce a new transgenic variety. Alternatively, a genetic traitwhich has been engineered into a particular canola variety using theforegoing transformation techniques could be moved into another varietyusing traditional backcrossing techniques that are well known in theplant breeding arts. For example, a backcrossing approach could be usedto move an engineered trait from a public, non-elite variety into anelite variety, or from a variety containing a foreign gene in its genomeinto a variety or varieties which do not contain that gene. As usedherein, “crossing” can refer to a simple X by Y cross, or the process ofbackcrossing, depending on the context.

Genetic Marker Profile Through SSR and First Generation Progeny

In addition to phenotypic observations, a plant can also be identifiedby its genotype. The genotype of a plant can be characterized through agenetic marker profile which can identify plants of the same variety ora related variety or be used to determine or validate a pedigree.Genetic marker profiles can be obtained by techniques such asRestriction Fragment Length Polymorphisms (RFLPs), Randomly AmplifiedPolymorphic DNAs (RAPDs), Arbitrarily Primed Polymerase Chain Reaction(AP-PCR), DNA Amplification Fingerprinting (DAF), Sequence CharacterizedAmplified Regions (SCARs), Amplified Fragment Length Polymorphisms(AFLPs), Simple Sequence Repeats (SSRs) which are also referred to asMicrosatellites, and Single Nucleotide Polymorphisms (SNPs).

Particular markers used for these purposes are not limited to anyparticular set of markers, but are envisioned to include any type ofmarker and marker profile which provides a means of distinguishingvarieties. One method of comparison is to use only homozygous loci forSCV528587.

In addition to being used for identification of canola variety SCV528587and plant parts and plant cells of variety SCV528587, the geneticprofile may be used to identify a canola plant produced through the useof SCV528587 or to verify a pedigree for progeny plants produced throughthe use of SCV528587. The genetic marker profile is also useful inbreeding and developing backcross conversions.

The present invention comprises a canola plant characterized bymolecular and physiological data obtained from the representative sampleof said variety deposited with the American Type Culture Collection(ATCC). Further provided by the invention is a canola plant formed bythe combination of the disclosed canola plant or plant cell with anothercanola plant or cell and comprising the homozygous alleles of thevariety.

Means of performing genetic marker profiles using SSR polymorphisms arewell known in the art. SSRs are genetic markers based on polymorphismsin repeated nucleotide sequences, such as microsatellites. A markersystem based on SSRs can be highly informative in linkage analysisrelative to other marker systems in that multiple alleles may bepresent. Another advantage of this type of marker is that, through useof flanking primers, detection of SSRs can be achieved, for example, bythe polymerase chain reaction (PCR), thereby eliminating the need forlabor-intensive Southern hybridization. The PCR detection is done by useof two oligonucleotide primers flanking the polymorphic segment ofrepetitive DNA. Repeated cycles of heat denaturation of the DNA followedby annealing of the primers to their complementary sequences at lowtemperatures, and extension of the annealed primers with DNA polymerase,comprise the major part of the methodology.

Following amplification, markers can be scored by electrophoresis of theamplification products. Scoring of marker genotype is based on the sizeof the amplified fragment, which may be measured by the number of basepairs of the fragment. While variation in the primer used or inlaboratory procedures can affect the reported fragment size, relativevalues should remain constant regardless of the specific primer orlaboratory used. When comparing varieties it is preferable if all SSRprofiles are performed in the same lab.

The SSR profile of canola plant SCV528587 can be used to identify plantscomprising SCV528587 as a parent, since such plants will comprise thesame homozygous alleles as SCV528587. Because the canola variety isessentially homozygous at all relevant loci, most loci should have onlyone type of allele present. In contrast, a genetic marker profile of anF₁ progeny should be the sum of those parents, e.g., if one parent washomozygous for allele x at a particular locus, and the other parenthomozygous for allele y at that locus, then the F₁ progeny will be xy(heterozygous) at that locus. Subsequent generations of progeny producedby selection and breeding are expected to be of genotype x (homozygous),y (homozygous), or xy (heterozygous) for that locus position. When theF₁ plant is selfed or sibbed for successive filial generations, thelocus should be either x or y for that position.

In addition, plants and plant parts substantially benefiting from theuse of SCV528587 in their development, such as SCV528587 comprising abackcross conversion, transgene, or genetic sterility factor, may beidentified by having a molecular marker profile with a high percentidentity to SCV528587. Such a percent identity might be 95%, 96%, 97%,98%, 99%, 99.5% or 99.9% identical to SCV528587.

The SSR profile of SCV528587 also can be used to identify essentiallyderived varieties and other progeny varieties developed from the use ofSCV528587, as well as cells and other plant parts thereof. Progenyplants and plant parts produced using SCV528587 may be identified byhaving a molecular marker profile of at least 25%, 30%, 35%, 40%, 45%,50%, 55%, 60%, 65%, 70%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, 99% or 99.5% genetic contribution from canola variety, as measuredby either percent identity or percent similarity. Such progeny may befurther characterized as being within a pedigree distance of SCV528587,such as within 1, 2, 3, 4 or 5 or less cross-pollinations to a canolaplant other than SCV528587 or a plant that has SCV528587 as aprogenitor. Unique molecular profiles may be identified with othermolecular tools such as SNPs and RFLPs.

While determining the SSR genetic marker profile of the plants describedsupra, several unique SSR profiles may also be identified which did notappear in either parent of such plant. Such unique SSR profiles mayarise during the breeding process from recombination or mutation. Acombination of several unique alleles provides a means of identifying aplant variety, an F₁ progeny produced from such variety, and progenyproduced from such variety.

Single-Gene Conversions

When the term “canola plant” is used in the context of the presentinvention, this also includes any single gene conversions of thatvariety. The term single gene converted plant as used herein refers tothose canola plants which are developed by backcrossing, whereinessentially all of the morphological and physiological characteristicsof a variety are recovered in addition to the single gene transferredinto the variety via the backcrossing technique. Backcrossing methodscan be used with the present invention to improve or introduce acharacteristic into the variety. A hybrid progeny may be backcrossed tothe recurrent parent 1, 2, 3, 4, 5, 6, 7, 8 or more times as part ofthis invention. The parental canola plant that contributes the gene forthe characteristic is termed the nonrecurrent or donor parent. Thisterminology refers to the fact that the nonrecurrent parent is used onetime in the backcross protocol and therefore does not recur. Theparental canola plant to which the gene or genes from the nonrecurrentparent are transferred is known as the recurrent parent as it is usedfor several rounds in the backcrossing protocol. In a typical backcrossprotocol, the original variety of interest (recurrent parent) is crossedto a second variety (nonrecurrent parent) that carries the single geneof interest to be transferred. The resulting progeny from this cross arethen crossed again to the recurrent parent and the process is repeateduntil a canola plant is obtained wherein essentially all of themorphological and physiological characteristics of the recurrent parentare recovered in the converted plant, in addition to the singletransferred gene from the nonrecurrent parent.

The selection of a suitable recurrent parent is an important step for asuccessful backcrossing procedure. The goal of a backcross protocol isto alter or substitute a single trait or characteristic in the originalvariety. To accomplish this, a single gene of the recurrent variety ismodified or substituted with the gene from the nonrecurrent parent,while retaining essentially all of the rest of the genetic, andtherefore the morphological and physiological, constitution of theoriginal variety. The choice of the particular nonrecurrent parent willdepend on the purpose of the backcross; one of the major purposes is toadd some agronomically important trait to the plant. The exactbackcrossing protocol will depend on the characteristic or trait beingaltered to determine an appropriate testing protocol. Althoughbackcrossing methods are simplified when the characteristic beingtransferred is a dominant allele, a recessive allele may also betransferred. In this instance it may be necessary to introduce a test ofthe progeny to determine if the characteristic has been successfullytransferred.

Many single gene traits have been identified that are not regularlyselected for in the development of a new variety but that can beimproved by backcrossing techniques. Single gene traits may or may notbe transgenic; examples of these traits include but are not limited to,male sterility, waxy starch, herbicide resistance, resistance forbacterial, fungal, or viral disease, insect resistance, male fertility,enhanced nutritional quality, industrial usage, yield stability andyield enhancement. These genes are generally inherited through thenucleus.

Introduction of a New Trait or Locus into SCV528587

Variety SCV528587 represents a new base genetic variety into which a newlocus or trait may be introgressed. Direct transformation andbackcrossing represent two methods that can be used to accomplish suchan introgression. The term backcross conversion and single locusconversion are used interchangeably to designate the product of abackcrossing program.

Backcross Conversions of SCV528587

A backcross conversion of SCV528587 may occur when DNA sequences areintroduced through backcrossing with SCV528587 utilized as the recurrentparent. Both naturally occurring and transgenic DNA sequences may beintroduced through backcrossing techniques. Molecular marker assistedbreeding or selection may be utilized to reduce the number ofbackcrosses necessary to achieve the backcross conversion.

The complexity of the backcross conversion method depends on the type oftrait being transferred (single genes or closely linked genes as vs.unlinked genes), the level of expression of the trait, the type ofinheritance (cytoplasmic or nuclear) and the types of parents includedin the cross. It is understood by those of ordinary skill in the artthat for single gene traits that are relatively easy to classify, thebackcross method is effective and relatively easy to manage. Traits thatmay be transferred through backcross conversion include, but are notlimited to, sterility (nuclear and cytoplasmic), fertility restoration,nutritional enhancements, drought tolerance, nitrogen utilization,altered fatty acid profile, altered seed amino acid levels, altered seedoil levels, low phytate, industrial enhancements, disease resistance(bacterial, fungal or viral), insect resistance and herbicideresistance. In addition, an introgression site itself, such as an FRTsite, Lox site or other site specific integration site, may be insertedby backcrossing and utilized for direct insertion of one or more genesof interest into a specific plant variety. In some embodiments of theinvention, the number of loci that may be backcrossed into SCV528587 isat least 1, 2, 3, 4, or 5 and/or no more than 6, 5, 4, 3, or 2. A singlelocus may contain several transgenes, such as a transgene for diseaseresistance that, in the same expression vector, also contains atransgene for herbicide resistance. The gene for herbicide resistancemay be used as a selectable marker and/or as a phenotypic trait. Asingle locus conversion of site specific integration system allows forthe integration of multiple genes at the converted loci.

The backcross conversion may result from either the transfer of adominant allele or a recessive allele. Selection of progeny containingthe trait of interest is accomplished by direct selection for a traitassociated with a dominant allele. Transgenes transferred viabackcrossing typically function as a dominant single gene trait and arerelatively easy to classify. Selection of progeny for a trait that istransferred via a recessive allele requires growing and selfing thefirst backcross generation to determine which plants carry the recessivealleles. Recessive traits may require additional progeny testing insuccessive backcross generations to determine the presence of the locusof interest. The last backcross generation is usually selfed to givepure breeding progeny for the gene(s) being transferred, although abackcross conversion with a stably introgressed trait may also bemaintained by further backcrossing to the recurrent parent withselection for the converted trait.

Along with selection for the trait of interest, progeny are selected forthe phenotype of the recurrent parent. The backcross is a form ofinbreeding, and the features of the recurrent parent are automaticallyrecovered after successive backcrosses. As noted above, the number ofbackcrosses necessary can be reduced with the use of molecular markers.Other factors, such as a genetically similar donor parent, may alsoreduce the number of backcrosses necessary. As noted, backcrossing iseasiest for simply inherited, dominant and easily recognized traits.

One process for adding or modifying a trait or locus in canola varietySCV528587 comprises crossing plants of canola variety SCV528587 grownfrom canola variety SCV528587 seed with plants of another canola varietythat comprise the trait or locus, selecting F₁ progeny plants thatcomprise the trait or locus to produce selected F₁ progeny plants,crossing the selected progeny plants with plants of canola SCV528587 toproduce backcross progeny plants, selecting for backcross progeny plantsthat have the trait or locus and the morphological characteristics ofcanola variety SCV528587 to produce selected backcross progeny plants;and backcrossing to SCV528587 three or more times in succession toproduce selected fourth or higher backcross progeny plants that comprisesaid trait or locus. The modified SCV528587 may be further characterizedas having essentially all of the morphological and physiologicalcharacteristics of canola variety SCV528587 listed in Table 1 and/or maybe characterized by percent similarity or identity to SCV528587 asdetermined by SSR markers. The above method may be utilized with fewerbackcrosses in appropriate situations, such as when the donor parent ishighly related or markers are used in the selection step. Traits thatmay be used include those nucleic acids known in the art, some of whichare listed herein, that will affect traits through nucleic acidexpression or inhibition. Loci that may be used include theintrogression of FRT, Lox and other sites for site specific integration,which may also affect a trait if a functional nucleic acid is insertedat the integration site.

In addition, the above process and other similar processes describedherein may be used to produce first generation progeny canola seed byadding a step at the end of the process that comprises crossingSCV528587 with the introgressed trait or locus with a different canolaplant and harvesting the resultant first generation progeny canola seed.

Tissue Culture of Canola

Further production of the SCV528587 variety can occur by tissue cultureand regeneration. Culture of various tissues of canola and regenerationof plants therefrom is known and widely published. Thus, another aspectof this invention is to provide cells which upon growth anddifferentiation produce canola plants having the physiological andmorphological characteristics of canola variety SCV528587.

As used herein, the term “tissue culture” indicates a compositioncomprising isolated cells of the same or a different type or acollection of such cells organized into parts of a plant. Exemplarytypes of tissue cultures are protoplasts, calli, plant clumps, and plantcells that can generate tissue culture that are intact in plants orparts of plants, such as embryos, pollen, flowers, seeds, pods, leaves,stems, roots, root tips, anthers, pistils and the like. Means forpreparing and maintaining plant tissue culture are well known in theart. Tissue culture comprising organs can be used in the presentinvention to produce regenerated plants.

Using SCV528587 to Develop Other Canola Varieties

Canola varieties such as SCV528587 are typically developed for use inseed and grain production. However, canola varieties such as SCV528587also provide a source of breeding material that may be used to developnew canola varieties. Plant breeding techniques known in the art andused in a canola plant breeding program include, but are not limited to,recurrent selection, mass selection, bulk selection, mass selection,backcrossing, pedigree breeding, open pollination breeding, restrictionfragment length polymorphism enhanced selection, genetic marker enhancedselection, making double haploids, and transformation. Oftencombinations of these techniques are used. The development of canolavarieties in a plant breeding program requires, in general, thedevelopment and evaluation of homozygous varieties.

Additional Breeding Methods

This invention is directed to methods for producing a canola plant bycrossing a first parent canola plant with a second parent canola plantwherein either the first or second parent canola plant is varietySCV528587. The other parent may be any other canola plant, such as acanola plant that is part of a synthetic or natural population. Any suchmethods using canola variety SCV528587 are part of this invention:selfing, sibbing, backcrosses, mass selection, pedigree breeding, bulkselection, hybrid production, crosses to populations, and the like.These methods are well known in the art and some of the more commonlyused breeding methods are described below.

The following describes breeding methods that may be used with canolavariety SCV528587 in the development of further canola plants. One suchembodiment is a method for developing a variety SCV528587 progeny canolaplant in a canola plant breeding program comprising: obtaining thecanola plant, or a part thereof, of variety SCV528587 utilizing saidplant or plant part as a source of breeding material and selecting acanola variety SCV528587 progeny plant with molecular markers in commonwith variety SCV528587 and/or with morphological and/or physiologicalcharacteristics selected from the characteristics listed in Table 1.Breeding steps that may be used in the canola plant breeding programinclude pedigree breeding, backcrossing, mutation breeding, andrecurrent selection. In conjunction with these steps, techniques such asRFLP-enhanced selection, genetic marker enhanced selection (for exampleSSR markers) and the making of double haploids may be utilized.

Another method involves producing a population of canola varietySCV528587 progeny canola plants, comprising crossing variety SCV528587with another canola plant, thereby producing a population of canolaplants, which, on average, derive 50% of their alleles from canolavariety SCV528587. A plant of this population may be selected andrepeatedly selfed or sibbed with a canola variety resulting from thesesuccessive filial generations. One embodiment of this invention is thecanola variety produced by this method and that has obtained at least50% of its alleles from canola variety SCV528587.

One of ordinary skill in the art of plant breeding would know how toevaluate the traits of two plant varieties to determine if there is nosignificant difference between the two traits expressed by thosevarieties. Thus the invention includes canola variety SCV528587 progenycanola plants comprising a combination of at least two variety SCV528587traits selected from the group consisting of those listed in Table 1 orthe variety SCV528587 combination of traits listed in the Summary of theInvention, so that said progeny canola plant is not significantlydifferent for said traits than canola variety SCV528587 as determined atthe 5% significance level when grown in the same environmentalconditions. Using techniques described herein, molecular markers may beused to identify said progeny plant as a canola variety SCV528587progeny plant. Mean trait values may be used to determine whether traitdifferences are significant, and preferably the traits are measured onplants grown under the same environmental conditions. Once such avariety is developed its value is substantial since it is important toadvance the germplasm base as a whole in order to maintain or improvetraits such as yield, disease resistance, pest resistance, and plantperformance in extreme environmental conditions.

Progeny of canola variety SCV528587 may also be characterized throughtheir filial relationship with canola variety SCV528587, as for example,being within a certain number of breeding crosses of canola varietySCV528587. A breeding cross is a cross made to introduce new geneticsinto the progeny, and is distinguished from a cross, such as a self or asib cross, made to select among existing genetic alleles. The lower thenumber of breeding crosses in the pedigree, the closer the relationshipbetween canola variety SCV528587 and its progeny. For example, progenyproduced by the methods described herein may be within 1, 2, 3, 4 or 5breeding crosses of canola variety SCV528587.

As used herein, the term “plant” includes plant cells, plantprotoplasts, plant cell tissue cultures from which canola plants can beregenerated, plant calli, plant clumps, and plant cells that are intactin plants or parts of plants, such as embryos, pollen, ovules, flowers,pods, leaves, roots, root tips, anthers, cotyledons, hypocotyls,meristematic cells, stems, pistils, petiole, and the like.

Pedigree Breeding

Pedigree breeding starts with the crossing of two genotypes, such asSCV528587 and another canola variety having one or more characteristicsthat is lacking or which complements SCV528587. If the two originalparents do not provide all the characteristics, other sources can beincluded in the breeding population. In the pedigree method, superiorplants are selfed and selected in successive filial generations. In thesucceeding filial generations the heterozygous condition gives way tohomogeneous varieties as a result of self-pollination and selection.Typically in the pedigree method of breeding, five or more successivefilial generations of selfing and selection is practiced: F₁ to F₂; F₂to F₃; F₃ to F₄; F₄ to F₅, etc. After a sufficient amount of inbreeding,successive filial generations will serve to increase seed of thedeveloped variety. Preferably, the developed variety compriseshomozygous alleles at about 95% or more of its loci.

In addition to being used to create a backcross conversion, backcrossingcan also be used in combination with pedigree breeding. As discussedpreviously, backcrossing can be used to transfer one or more traits fromone variety, the donor parent, to a developed variety called therecurrent parent, which has overall good agronomic characteristics yetlacks that trait or traits. However, the same procedure can be used tomove the progeny toward the genotype of the recurrent parent but at thesame time retain many components of the non-recurrent parent by stoppingthe backcrossing at an early stage and proceeding with selfing andselection. For example, a canola variety may be crossed with anothervariety to produce a first generation progeny plant. The firstgeneration progeny plant may then be backcrossed to one of its parentvarieties to create a BC1 or BC2. Progeny are selfed and selected sothat the newly developed variety has many of the attributes of therecurrent parent and yet several of the attributes of the non-recurrentparent. This approach leverages the value and strengths of the recurrentparent for use in new canola varieties.

Therefore, an embodiment of this invention is a method of making abackcross conversion of canola variety SCV528587, comprising the stepsof crossing a plant of canola variety SCV528587 with a donor plantcomprising a trait, selecting an F₁ progeny plant comprising the trait,and backcrossing the selected F₁ progeny plant to a plant of canolavariety SCV528587. This method may further comprise the step ofobtaining a molecular marker profile of canola variety SCV528587 andusing the molecular marker profile to select for a progeny plant withthe trait and the molecular marker profile of SCV528587. In oneembodiment the trait is a mutant gene or transgene present in the donorparent.

Recurrent Selection and Mass Selection

Recurrent selection is a method used in a plant breeding program toimprove a population of plants. SCV528587 is suitable for use in arecurrent selection program. The method entails individual plants crosspollinating with each other to form progeny. The progeny are grown andthe superior progeny selected by any number of selection methods, whichinclude individual plant, half-sib progeny, full-sib progeny and selfedprogeny. The selected progeny are cross-pollinated with each other toform progeny for another population. This population is planted andagain superior plants are selected to cross-pollinate with each other.Recurrent selection is a cyclical process and therefore can be repeatedas many times as desired. The objective of recurrent selection is toimprove the traits of a population. The improved population can then beused as a source of breeding material to obtain new varieties forcommercial or breeding use, including the production of a syntheticvariety. A synthetic variety is the resultant progeny formed by theintercrossing of several selected varieties.

Mass selection is a useful technique when used in conjunction withmolecular marker enhanced selection. In mass selection seeds fromindividuals are selected based on phenotype or genotype. These selectedseeds are then bulked and used to grow the next generation. Bulkselection requires growing a population of plants in a bulk plot,allowing the plants to self-pollinate, harvesting the seed in bulk andthen using a sample of the seed harvested in bulk to plant the nextgeneration. Also, instead of self-pollination, directed pollinationcould be used as part of the breeding program.

Mutation Breeding

Mutation breeding is another method of introducing new traits intocanola variety SCV528587. Mutations that occur spontaneously or areartificially induced can be useful sources of variability for a plantbreeder. The goal of artificial mutagenesis is to increase the rate ofmutation for a characteristic. Mutation rates can be increased by manydifferent means including temperature, long-term seed storage, tissueculture conditions, radiation; such as X-rays, Gamma rays (e.g. cobalt60 or cesium 137), neutrons, (product of nuclear fission by uranium 235in an atomic reactor), Beta radiation (emitted from radioisotopes suchas phosphorus 32 or carbon 14), or ultraviolet radiation (preferablyfrom 2500 to 2900 nm), or chemical mutagens (such as base analogues(5-bromo-uracil), related compounds (8-ethoxy caffeine), antibiotics(streptonigrin), alkylating agents (sulfur mustards, nitrogen mustards,epoxides, ethylenamines, sulfates, sulfonates, sulfones, lactones),azide, hydroxylamine, nitrous acid, or acridines. Once a trait isobserved through mutagenesis the trait may then be incorporated intoexisting germplasm by traditional breeding techniques. In addition,mutations created in other canola plants may be used to produce abackcross conversion of canola variety SCV528587 that comprises suchmutation.

Breeding with Molecular Markers

Molecular markers, which include markers identified through the use oftechniques such as Isozyme Electrophoresis, Restriction Fragment LengthPolymorphisms (RFLPs), Randomly Amplified Polymorphic DNAs (RAPDs),Arbitrarily Primed Polymerase Chain Reaction (AP-PCR), DNA AmplificationFingerprinting (DAF), Sequence Characterized Amplified Regions (SCARs),Amplified Fragment Length Polymorphisms (AFLPs), Simple Sequence Repeats(SSRs) and Single Nucleotide Polymorphisms (SNPs), may be used in plantbreeding methods utilizing canola variety SCV528587. One use ofmolecular markers is Quantitative Trait Loci (QTL) mapping. QTL mappingis the use of markers, which are known to be closely linked to allelesthat have measurable effects on a quantitative trait. Selection in thebreeding process is based upon the accumulation of markers linked to thepositive effecting alleles and/or the elimination of the markers linkedto the negative effecting alleles from the plant's genome.

Molecular markers can also be used during the breeding process for theselection of qualitative traits. For example, markers closely linked toalleles or markers containing sequences within the actual alleles ofinterest can be used to select plants that contain the alleles ofinterest during a backcrossing breeding program. The markers can also beused to select for the genome of the recurrent parent and against thegenome of the donor parent. Using this procedure can minimize the amountof genome from the donor parent that remains in the selected plants. Itcan also be used to reduce the number of crosses back to the recurrentparent needed in a backcrossing program. The use of molecular markers inthe selection process is often called genetic marker enhanced selection.Molecular markers may also be used to identify and exclude certainsources of germplasm as parental varieties or ancestors of a plant byproviding a means of tracking genetic profiles through crosses.

Production of Double Haploids

The production of double haploids can also be used for the developmentof plants with a homozygous phenotype in the breeding program. Forexample, a canola plant for which canola variety SCV528587 is a parentcan be used to produce double haploid plants. Double haploids areproduced by the doubling of a set of chromosomes (1 N) from aheterozygous plant to produce a completely homozygous individual. Thiscan be advantageous because the process omits the generations of selfingneeded to obtain a homozygous plant from a heterozygous source.

Haploid induction systems have been developed for various plants toproduce haploid tissues, plants and seeds. Thus, an embodiment of thisinvention is a process for making a substantially homozygous SCV528587progeny plant by producing or obtaining a seed from the cross ofSCV528587 and another canola plant and applying double haploid methodsto the F₁ seed or F₁ plant or to any successive filial generation. Basedon studies in maize and currently being conducted in canola, suchmethods would decrease the number of generations required to produce avariety with similar genetics or characteristics to SCV528587.

In particular, a process of making seed retaining the molecular markerprofile of canola variety SCV528587 is contemplated, such processcomprising obtaining or producing F₁ seed for which canola varietySCV528587 is a parent, inducing doubled haploids to create progenywithout the occurrence of meiotic segregation, obtaining the molecularmarker profile of canola variety SCV528587, and selecting progeny thatretain the molecular marker profile of SCV528587.

A pollination control system and effective transfer of pollen from oneparent to the other offers improved plant breeding and an effectivemethod for producing hybrid canola seed and plants. For example, theogura cytoplasmic male sterility (cms) system, developed via protoplastfusion between radish (Raphanus sativus) and rapeseed (Brassica napus)is one of the most frequently used methods of hybrid production. Itprovides stable expression of the male sterility trait and an effectivenuclear restorer gene.

In developing improved new Brassica hybrid varieties, breeders useself-incompatible (SI), cytoplasmic male sterile (CMS) and nuclear malesterile (NMS) Brassica plants as the female parent. In using theseplants, breeders are attempting to improve the efficiency of seedproduction and the quality of the F₁ hybrids and to reduce the breedingcosts. When hybridization is conducted without using SI, CMS or NMSplants, it is more difficult to obtain and isolate the traits in theprogeny (F₁ generation) because the parents are capable of undergoingboth cross-pollination and self-pollination. If one of the parents is aSI, CMS or NMS plant that is incapable of producing pollen, onlycross-pollination will occur. By eliminating the pollen of one parentalvariety in a cross, a plant breeder is assured of obtaining hybrid seedof uniform quality, provided that the parents are of uniform quality andthe breeder conducts a single cross.

In one instance, production of F₁ hybrids includes crossing a CMSBrassica female parent, with a pollen producing male Brassica parent. Toreproduce effectively, however, the male parent of the F₁ hybrid musthave a fertility restorer gene (Rf gene). The presence of an Rf genemeans that the F₁ generation will not be completely or partiallysterile, so that either self-pollination or cross-pollination may occur.Self-pollination of the F₁ generation to produce several subsequentgenerations is important to ensure that a trait is heritable and stableand that a new variety has been isolated.

An example of a Brassica plant which is cytoplasmic male sterile andused for breeding is ogura (OGU) cytoplasmic male sterile. A fertilityrestorer for ogura cytoplasmic male sterile plants has been transferredfrom Raphanus sativus (radish) to Brassica. The restorer gene is Rf1,originating from radish. Improved versions of this restorer have beendeveloped as well. Other sources and refinements of CMS sterility incanola include the Polima cytoplasmic male sterile plant.

Further, as a result of the advances in sterility systems, varieties aredeveloped that can be used as an open pollinated line (i.e. a pure linesold to the grower for planting) and/or as a sterile inbred (female)used in the production of F₁ hybrid seed. In the latter case, favorablecombining ability with a restorer (male) would be desirable. Theresulting hybrid seed would then be sold to the grower for planting.

The development of a canola hybrid in a canola plant breeding programinvolves three steps: (1) the selection of plants from various germplasmpools for initial breeding crosses; (2) the selfing of the selectedplants from the breeding crosses for several generations to produce aseries of inbred lines, which, although different from each other, breedtrue and are highly uniform; and (3) crossing the selected inbred lineswith different inbred lines to produce the hybrids. During theinbreeding process in canola, the vigor of the lines decreases. Vigor isrestored when two different inbred lines are crossed to produce thehybrid. An important consequence of the homozygosity and homogeneity ofthe inbred lines is that the hybrid between a defined pair of inbredswill always be the same. Once the inbreds that give a superior hybridhave been identified, the hybrid seed can be reproduced indefinitely aslong as the homogeneity of the inbred parents is maintained.

Combining ability of a line, as well as the performance of the line perse, is a factor in the selection of improved canola varieties that maybe used as inbreds. Combining ability refers to a line's contribution asa parent when crossed with other lines to form hybrids. The hybridsformed for the purpose of selecting superior lines are designated testcrosses. One way of measuring combining ability is by using breedingvalues. Breeding values are based on the overall mean of a number oftest crosses. This mean is then adjusted to remove environmental effectsand is adjusted for known genetic relationships among the lines.

Hybrid seed production requires inactivation of pollen produced by thefemale parent. Incomplete inactivation of the pollen provides thepotential for self-pollination. This inadvertently self-pollinated seedmay be unintentionally harvested and packaged with hybrid seed.Similarly, because the male parent is grown next to the female parent inthe field, there is also the potential that the male selfed seed couldbe unintentionally harvested and packaged with the hybrid seed. Once theseed from the hybrid bag is planted, it is possible to identify andselect these self-pollinated plants. These self-pollinated plants willbe genetically equivalent to one of the inbred lines used to produce thehybrid. Though the possibility of inbreds being included in hybrid seedbags exists, the occurrence is rare because much care is taken to avoidsuch inclusions. These self-pollinated plants can be identified andselected by one skilled in the art, either through visual or molecularmethods.

Brassica napus canola plants, absent the use of sterility systems, arerecognized to commonly be self-fertile with approximately 70 to 90percent of the seed normally forming as the result of self-pollination.The percentage of cross-pollination may be further enhanced whenpopulations of recognized insect pollinators at a given growing site aregreater. Thus open pollination is often used in commercial canolaproduction.

Industrial Uses

Currently Brassica napus canola is recognized as an increasinglyimportant oilseed crop and a source of meal in many parts of the world.The oil as removed from the seeds commonly contains a lesserconcentration of endogenously formed saturated fatty acids than othervegetable oils and is well suited for use in the production of salad oilor other food products or in cooking or frying applications. The oilalso finds utility in industrial applications. Additionally, the mealcomponent of the seeds can be used as a nutritious protein concentratefor livestock.

Canola oil has the lowest level of saturated fatty acids of allvegetable oils. “Canola” refers to rapeseed (Brassica) which has aerucic acid (C22:1) content of at most 2 percent by weight based on thetotal fatty acid content of a seed, and which produces, after crushing,an air-dried meal containing less than 30 micromoles (μmol) per gram ofdefatted (oil-free) meal. These types of rapeseed are distinguished bytheir edibility in comparison to more traditional varieties of thespecies.

Canola variety SCV528587 can be used in the production of an ediblevegetable oil or other food products in accordance with knowntechniques. The solid meal component derived from seeds can be used as anutritious livestock feed. Parts of the plant not used for human oranimal food can be used for biofuel.

Tables

In Table 2, selected oil quality characteristics of the seed of canolavariety SCV528587 are compared with oil quality characteristics of twoproprietary canola varieties. The data in Table 2 includes results onseed samples collected from two testing locations and are presented asaverages of the values observed. Column 1 shows the variety, column 2shows the percent saturated fatty acid content, column 3 shows thepercent oleic acid content, column 4 shows the percent linoleic contentand column 5 shows the percent linolenic content. Characteristics of ahybrid containing SCV528587 as a parent compared to two commercialvarieties are set forth in Table 3.

TABLE 2 Oil Quality Characteristics of SCV528587 Compared to TwoProprietary Canola Varieties % Sat. Fat. % Oleic Variety Acid Acid %Linoleic % Linolenic SCV528587 6.76 62.91 20.24 7.86 SCV318181 7.0059.45 21.53 9.64 SCV167385 6.57 60.36 22.72 7.95

TABLE 3 Characteristics of Hybrid G42437, Containing SCV528587, Comparedto Two Commercial Varieties* Yield Lodging DMat Sats Height Gluc OilProt BL FW CR Variety % 1 = best Days % cm μm/g % % rating rating rating45H29 98.8% 94.1 6.68 127 12.49 50.0 43.6 R R R 5440 101.2% 1.6 94.96.51 133 10.34 47.7 43.3 R R S Avg of Checks 100.0% 94.5 6.60 130 11.4248.8 43.4 G42437 98.0% 2.5 94.2 6.48 122 9.99 49.4 45.6 R R R #locations 40 9 36 27 20 27 27 27 10 1 1

DEPOSIT INFORMATION

A deposit of the canola variety SCV528587, which is disclosed hereinabove and referenced in the claims, will be made with the American TypeCulture Collection (ATCC), 10801 University Blvd., Manassas, Va.20110-2209. The date of deposit is ______ and the accession number forthose deposited seeds of canola variety SCV528587 is ATCC Accession No.______. All restrictions upon the deposit have been removed, and thedeposit is intended to meet all of the requirements of the BudapestTreaty and 37 C.F.R. § 1.801-1.809. The deposit will be maintained inthe depository for a period of 30 years, or 5 years after the lastrequest, or for the effective life of the patent, whichever is longer,and will be replaced if necessary during that period.

While a number of exemplary aspects and embodiments have been discussedabove, those of skill in the art will recognize certain modifications,permutations, additions and sub-combinations thereof. It is thereforeintended that the following appended claims and claims hereafterintroduced are interpreted to include all such modifications,permutations, additions and sub-combinations as are within their truespirit and scope.

1. A seed of canola variety SCV528587, wherein representative seed ofsaid canola variety have been deposited under ATCC Accession No. ______.2. A plant of canola variety SCV528587 or a part thereof, wherein theplant of canola variety SCV528587 is produced by growing the seed ofclaim 1; and wherein the plant part comprises at least one cell of theplant of canola variety SCV528587.
 3. A tissue culture produced fromprotoplasts, spheroplasts, or cells from the plant of claim 2, whereinat least one of the cells, spheroplasts, or protoplasts of the tissueculture are produced from a plant part selected from the groupconsisting of leaf, pollen, embryo, cotyledon, hypocotyl, meristematiccell, root, root tip, anther, pistil, flower, shoot, stem, petiole, andpod.
 4. A canola plant regenerated from the tissue culture of claim 3,wherein the regenerated plant comprises all of the morphological andphysiological characteristics of a plant of canola variety SCV528587. 5.A composition comprising a cultivation medium and the seed of claim 1 ora plant part of a canola plant grown from the seed, and wherein theplant part comprises at least one cell of the canola plant.
 6. Thecomposition of claim 5, wherein the cultivation medium is soil or asynthetic medium.
 7. A canola seed, wherein the canola seed is producedby crossing the plant of claim 2 with itself or a second canola plant.8. A canola plant or a part thereof, wherein the canola plant isproduced by growing the seed of claim 7; and wherein the plant partcomprises at least one cell of the canola plant.
 9. A method ofproducing a male sterile canola plant, the method comprising crossingthe plant of claim 2 with a male sterile canola plant, harvesting theresultant seed, growing the resultant seed, and selecting a male sterileplant grown form the resultant seed.
 10. A plant of canola varietySCV528587 further comprising a transgene, wherein the transgene wasintroduced into the plant by backcrossing or genetic transformation,wherein said plant otherwise comprises all of the morphological andphysiological characteristics of said canola variety, and whereinrepresentative seed of said canola variety have been deposited underATCC Accession No. ______.
 11. A plant of canola variety SCV528587further comprising a single locus conversion, wherein said plantotherwise comprises all of the morphological and physiologicalcharacteristics of said canola variety, and wherein representative seedof said canola variety have been deposited under ATCC Accession No.______.
 12. The plant of claim 11, wherein the single locus conversioncomprises a transgene.
 13. The plant of claim 11, wherein the singlelocus conversion comprises a nucleic acid sequence that encodes asite-specific recombinase or confers a trait selected from the groupconsisting of male sterility, herbicide tolerance, insect or pestresistance, disease resistance, modified fatty acid metabolism, andmodified carbohydrate metabolism.
 14. The plant of claim 11, wherein thesingle locus conversion confers tolerance to an herbicide selected fromthe group consisting of benzonitrile herbicides, cyclohexanedioneherbicides, imidazolinone herbicides, phenoxy herbicides, sulfonylureaherbicides, triazine herbicides, 1-aminocyclopropane-1-carboxylic acidsynthase-inhibiting herbicides, 4-hydroxyphenylpyruvatedioxygenase-inhibiting herbicides, acetolactate synthase-inhibitingherbicides, protoporphyrinogen oxidase-inhibiting herbicides,2,4-dichlorophenoxyacetic acid (2,4-D), bromoxynil, dicamba,glufosinate, glyphosate, nicosulfuron, and quizalofop-p-ethyl or encodesa protein selected from the group consisting of Bacillus thuringiensisendotoxin, fructosyltransferase, levansucrase, α-amylase, invertase, andstarch branching enzyme, or encodes an antisense nucleic acid of astearyl-ACP desaturase gene.
 15. A method of producing an industrialplant product, said method comprising obtaining the seed of claim 1 andproducing the industrial plant product therefrom.
 16. The method ofclaim 15, wherein the industrial plant product is selected from thegroup consisting of canola meal, livestock feed, protein concentrate,unblended canola oil, salad oil, cooking oil, frying oil, vegetable oil,a blended oil, and biofuel.
 17. A method of plant breeding, the methodcomprising the steps of: (a) crossing the plant of claim 2 with a secondcanola plant that comprises a heritable trait to produce an F₁ progenyplant that comprises the heritable trait; (b) selecting the F₁ progenyplant that comprises the heritable trait to produce a selected progenyplant; (c) crossing the selected progeny plant from step (b) with aplant of canola variety SCV528587 to produce a progeny plant of asubsequent generation that comprises the heritable trait; and (d)repeating steps (b) and (c) with the progeny plant of the subsequentgeneration produced from step (c) used in place of the F₁ progeny plantof step (b) during the repeating, wherein steps (b) and (c) are repeateduntil at least a backcross progeny plant is produced comprising theheritable trait.
 18. A plant produced by the method of claim 17, whereinthe plant comprises the heritable trait; and wherein said plantotherwise comprises all of the morphological and physiologicalcharacteristics of a plant of canola variety SCV528587.
 19. The plant ofclaim 18, wherein the heritable trait is herbicide resistance and theresistance is conferred to an herbicide selected from the groupconsisting of benzonitrile herbicides, cyclohexanedione herbicides,imidazolinone herbicides, phenoxy herbicides, sulfonylurea herbicides,triazine herbicides, 1-aminocyclopropane-1-carboxylic acidsynthase-inhibiting herbicides, 4-hydroxyphenylpyruvatedioxygenase-inhibiting herbicides, acetolactate synthase-inhibitingherbicides, protoporphyrinogen oxidase-inhibiting herbicides,2,4-dichlorophenoxyacetic acid (2,4-D), bromoxynil, dicamba,glufosinate, glyphosate, nicosulfuron, and quizalofop-p-ethyl.
 20. Theplant of claim 18, wherein the heritable trait is insect resistance,modified fatty acid metabolism, or modified carbohydrate metabolism andsaid heritable trait is conferred by a nucleic acid encoding a proteinselected from the group consisting of Bacillus thuringiensis endotoxin,phytase, fructosyltransferase, levansucrase, α-amylase, invertase, andstarch branching enzyme, or encoding an antisense nucleic acid of astearyl-ACP desaturase gene.